Functional chronic pain syndromes have no known pathophysiological basis yet affect millions of people in the United States. Many of these syndromes present as comorbid conditions. As many as 60% of patients with temporomandibular disorder (TMD) also report symptoms of abdominal pain consistent with irritable bowel syndrome (IBS), but the underlying connection between these comorbid conditions is unclear. There are some common clinical features, namely both conditions are substantially more common in women, the level of pain fluctuates during the menstrual cycle and they may be triggered or exacerbated by stress, but a mechanism that can connect these two conditions is unknown. The current application proposes a new animal model of comorbid pain conditions in which TMD and subchronic stress are combined to produce colonic hypersensitivity arising from otherwise healthy tissue. The longterm hypothesis is that persistent pain combined with stress alters the endogenous analgesia system resulting in enhanced secondary hyperalgesia in close proximity to the initial painful site, but also in hypersensitivityin distal deep tissue. The current application addresses the hypothesis that masseter muscle inflammation plus stress induces colonic hypersensitivity that is modulated by gonadal hormones. Furthermore, the visceral hypersensitivity results from central sensitization at the level of the spinal cord. This will be tested in two specific aims: Specific aim 1: Characterize a new model of comorbid pain conditions in which visceral hypersensitivity is induced by a combination of masseter muscle inflammation and subchronic stress. TMD and IBS are individually more prevalent in women and are sensitive to hormonal fluctuation during the menstrual cycle. In these individual animal models (masseter muscle inflammation, colorectal distention), females are more sensitive than males and estrogen increases sensitivity in ovariectomized rats. This aim will characterize the effects of estradiol on visceral and somatic sensitivity in the comorbidity model and test the hypothesis that the visceral hypersensitivity induced by the comorbid conditions is modulated by estradiol. Specific Aim 2: Determine if the visceral hypersensitivity in the comorbid pain model results from sensitization in the periphery or at the level of the spinal cord. As a first step in determining the underlying mechanisms that account for the increase in visceral sensitivity following injury plus stress we will determine if there are increases in neurochemical markers indicative of peripheral or central sensitization. Such changes will indicate that injury plus stress activates either a descending pathway or hormonal surge that sensitizes primary afferents and/or dorsal horn neurons. Failure to observe any neurochemical upregulation will support an alternative hypothesis that the visceral hypersensitivity results from sensitization of nociceptive processing above the level of the spinal cord. Either result will guide future studies into mechanisms underlying comorbid pain conditions.